MOLECULAR ANALYSIS OF A MUSCLE ACTIN-CAPPING PROTEIN

肌动蛋白是一种普遍存在的蛋白质，参与许多结构和功能， 各种细胞中的运动过程。 我们假设蛋白质 与肌动蛋白丝末端结合的蛋白质在形成中很重要， 肌动蛋白丝与其他结构的定向和附着。 到 为了探索这一假说，我们从鸡骨骼中分离了一种蛋白质， 与肌动蛋白的倒刺末端有高亲和力结合的肌肉（Cap Z） 纤维，并定位到Z线。 Cap Z对肌动蛋白和 它在肌肉中位置表明CapZ可能在形成中起重要作用 以及肌动蛋白丝与Z线的连接。 我们最近 鉴定的cDNA编码的蛋白质是同源的两个亚基， Cap Z（两个用于α亚基，一个用于β亚基）。其他研究 显示Cap Z与来自网骨藻的蛋白高度同源， Xenopus laevis和C.在生物化学上不同于其他的 肌动蛋白帽蛋白 其中一种蛋白质（来自非洲爪蟾）具有 核定位 也有人提出，Cap Z与 一种与肌动蛋白丝的相对端或尖端结合的蛋白质 而Cap Z本身可能有核定位。 这个目标 该项目旨在确定鸡Cap Z的生物活性， 同源物 研究Cap Z在胚胎发生过程中的表达 使用北方和西方印迹来观察是否存在差异表达 同源亚基的结构 Cap Z的亚基将在 体外使用细菌表达系统及其对肌动蛋白和 将详细检查DNA结合特性。 表达的 蛋白质将被用来产生识别每个亚基的抗体 特别是用于研究它们的表达和组织定位。 我们还将通过产生一系列的 缺失或插入突变蛋白质和研究蛋白水解切割 Cap Z的片段 这些研究应该提供重要的信息 关于肌动蛋白帽蛋白的结构和功能， 与一些基本进程的相关性， 正常骨骼肌和心肌的细胞运动和相关的 涉及肌动蛋白的细胞过程。

Actin is a ubiquitous protein that is involved in numerous structural and motile processes in a variety of cells. We have postulated that proteins that bind to the ends of actin filaments are important in the formation, orientation and attachment of actin filaments to other structures. To explore this hypothesis, we have isolated a protein from chicken skeletal muscle (Cap Z) that binds with high affinity to the barbed end of actin filaments and is localized to the Z-line. The effect of Cap Z on actin and its location in muscle suggest that Cap Z may be important in the formation and attachment of actin filaments to the Z-line. We have recently identified cDNAs encoding proteins that are homologous to both subunits of Cap Z (two for the alpha subunit, one for the beta) . Other studies have shown that Cap Z is highly homologous to proteins from Dictyostelium, Xenopus laevis and C. elegans that are biochemically distinct from other actin-capping proteins. One of these proteins (from Xenopus laevis) has a nuclear localization. It has also been suggested that Cap Z is related to a protein that binds to the opposite, or pointed, end of actin filaments and that Cap Z itself may have a nuclear localization. The goal of this project is to define the biologic activities of the chicken Cap Z homologues. The expression of Cap Z during embryogenesis will be studied using Northern and Western blots to see if there is differential expression of the homologous subunits. The subunits of Cap Z will be expressed in vitro using bacterial expression systems and their effect on actin and DNA-binding characteristics will be examined in detail. The expressed proteins will be used to generate antibodies that recognize each subunit specifically for use in studying their expression and tissue localization. We will also determine the binding site for actin by generating a series of deletion or insertion mutant proteins and studying proteolytic cleavage fragments of Cap Z. These studies should provide important information about the structure and function of actin-capping proteins that will be of relevance to a number of fundamental processes, including the development of normal skeletal and cardiac muscle, cell motility and the related cellular processes that involve actin.